We recently showed that each of the yeast (S. cerevisiae) genes UBI1-UBI3 encodes a hybrid protein whose cleavage yields ubiquitin and a ribosomal protein[1]. The transient covalent association of ubiquitin and the tail within the UBI3 protein was shown to facilitate ribosome biogenesis, apparently by increasing the efficiency of the tail's incorporation into nascent ribosomes. While ubiquitin has long been known as a post - translational modifying group which signals the degradation of acceptor proteins[2-7], our results suggest a novel, "chaperone" function for ubiquitin, in which its covalent association with other proteins can promote the formation of specific cellular structures. We will test whether this chaperone function of ubiquitin operates through transient metabolic stabilization of the newly-formed tail, or by increasing the rate of the tail's transport to and assembly within nascent ribosomes. Techniques that allow kinetic analysis of the fate of newly-synthesized ubiquitin-tail hybrids will be developed and used to determine the rates of deubiquitination, incorporation within the ribosome, and degradation of tail proteins expressed either in wild-type cells or from appropriate mutants and overproducing strains. Similar experiments will test the applicability of our conclusions to genes which encode ubiquitin-like domains linked to non-ubiquitin domains [8-12]. Additional aims include: 1) The degradation of a variety of proteins apparently requires that ubiquitin moieties be attached sequentially to an acceptor protein to form a chain of branched ubiquitin-ubiquitin conjugates in which the carboxyl terminus of one ubiquitin is joined to Lys-48 of an adjacent ubiquitin; mutant ubiquitin molecules carrying Arg-48 can be conjugated to different protein but not to each other[13]. The function of ubiquitin-ubiquitin conjugation will be investigated using yeast strains in which all natural ubiquitin-coding elements are deleted and the Arg-48 mutant serves as the sole ubiquitin species. 2) The systematic isolation and analysis of yeast genes encoding proteins that are posttranslationally ubiquitinated. 3) Analysis of the role of the 15-kD interferon-induced homologue of ubiquitin[9-11] in viral infection. These sturdies should elucidate specific mechanisms whereby the attachment of ubiquitin to proteins regulates their function. Such insights may provide a foundation for understanding the poorly-defined roles of ubiquitin in DNA repair[14], cell cycle control[13-15], stress responses[3,16-22], neurodegenerative disease[23-25], cell-surface receptor function[26-28], and the turnover of short-lived proteins[2-7,13].